Copper deficiency results in the intracellular accumulation of iron. This iron is associated with the mitochondrial fraction and is thought to accumulate because of impaired mitochondrial uptake and subsequent heme synthesis. The purposes of this study are to define the nature of the accumulated iron and to use this iron material to identify more clearly the specific steps which are required to move iron across the mitochondrial membrane and ultimately into the heme molecule. Initial studies of the distribution of iron accumulation within the various organs and subcellular fractions of copper deficient rats have shown that at low doses of iron loading, iron accumulates within the mitochondrial fraction. Further studies will be directed toward localization of this iron by additional mitochondrial fractionation. Initial characterization of the accumulated iron shows it to reside in a material which is excluded by Sepharose 4B. This material will support mitochondrial heme synthesis. Proposed studies will be directed toward the characterization of this material as well as the use of it for substrate to define the steps required for mitochondrial iron uptake and heme synthesis.